The ATP-binding cassette transporter subfamily G member 4 mediates cuticular hydrocarbon transport to regulate drought tolerance in Acyrthosiphon pisum.

ABC transporters are a highly conserved membrane protein class that promote the transport of substances across membranes. Under drought conditions, insects primarily regulate the content of cuticular hydrocarbon (CHC) to retain water and prevent evaporative loss. Involvement of ABC transporter protein G (ABCG) subfamily genes in insect CHC transport has been relatively understudied. In this study, we demonstrated that ABCG4 gene in Acyrthosiphon pisum (ApABCG4) is involved in CHC transport and affects drought tolerance by regulating CHC accumulation. ApABCG4 is strongly expressed in the abdominal cuticle and embryonic stages of A. pisum. Effective silencing of ApABCG4 was achieved using RNAi, and the silencing duration was analyzed. ApABCG4 silencing resulted in a significant decrease in the total and component contents of the CHC and cuticular waxy coatings of A. pisum. Nevertheless, the internal hydrocarbon content remained unchanged. The lack of cuticular hydrocarbons significantly reduced the drought tolerance of A. pisum, shortening its survival time under drought stress. Drought stress caused significant upregulation of ApABCG4. Molecular docking showed that ApABCG4 has a high binding affinity for nine n-alkanes of CHC through electrostatic interactions. These results indicate that ApABCG4 is a novel RNAi target with key applications in aphid biological control.

Oxygen-carrying semiconducting polymer nanoprodrugs induce sono-pyroptosis for deep-tissue tumor treatment.

Sonodynamic therapy (SDT) has been explored for cancer therapy, especially for deep tumors due to its low tissue penetration restriction. The therapeutic efficacy of SDT is limited due to the complicated tumor microenvironment. This study reports the construction of oxygen-carrying semiconducting polymer nanoprodrugs (OSPNpro) for deep tumor treatment via combining amplified SDT with pyroptosis. An oxygen carrier perfluorohexane, sonodynamic semiconducting polymer as the sonosensitizer, and reactive oxygen species (ROS)-responsive prodrug are co-loaded into a nanoparticle system, leading to the formation of these polymer nanoprodrugs. Such OSPNpro show an effective accumulation in tumor tissues after systemic administration, in which they deliver oxygen to relieve tumor hypoxia microenvironment and thus mediate amplified SDT via producing ROS under ultrasound (US) irradiation, even when the tumors are covered with a 2-cm chicken breast tissue. In addition, the ROS-responsive prodrugs are activated by the generated ROS to trigger pyroptosis of tumor cells. Such a sono-pyroptosis induces a strong antitumor immunity with obviously higher level infiltrations of effector immune cells into tumors. Therefore, OSPNpro-based combinational therapy can greatly inhibit the growth of 2-cm chicken breast tissue-covered deep tumors and suppress tumor metastasis. This study offers a prodrug nanoplatform for treatment of deep tumor via sono-pyroptosis strategy.

Tree-based ensemble machine learning models in the prediction of acute respiratory distress syndrome following cardiac surgery: a multicenter cohort study.

BACKGROUND: Acute respiratory distress syndrome (ARDS) after cardiac surgery is a severe respiratory complication with high mortality and morbidity. Traditional clinical approaches may lead to under recognition of this heterogeneous syndrome, potentially resulting in diagnosis delay. This study aims to develop and external validate seven machine learning (ML) models, trained on electronic health records data, for predicting ARDS after cardiac surgery. METHODS: This multicenter, observational cohort study included patients who underwent cardiac surgery in the training and testing cohorts (data from Nanjing First Hospital), as well as those patients who had cardiac surgery in a validation cohort (data from Shanghai General Hospital). The number of important features was determined using the sliding windows sequential forward feature selection method (SWSFS). We developed a set of tree-based ML models, including Decision Tree, GBDT, AdaBoost, XGBoost, LightGBM, Random Forest, and Deep Forest. Model performance was evaluated using the area under the receiver operating characteristic curve (AUC) and Brier score. The SHapley Additive exPlanation (SHAP) techinque was employed to interpret the ML model. Furthermore, a comparison was made between the ML models and traditional scoring systems. ARDS is defined according to the Berlin definition. RESULTS: A total of 1996 patients who had cardiac surgery were included in the study. The top five important features identified by the SWSFS were chronic obstructive pulmonary disease, preoperative albumin, central venous pressure_T4, cardiopulmonary bypass time, and left ventricular ejection fraction. Among the seven ML models, Deep Forest demonstrated the best performance, with an AUC of 0.882 and a Brier score of 0.809 in the validation cohort. Notably, the SHAP values effectively illustrated the contribution of the 13 features attributed to the model output and the individual feature's effect on model prediction. In addition, the ensemble ML models demonstrated better performance than the other six traditional scoring systems. CONCLUSIONS: Our study identified 13 important features and provided multiple ML models to enhance the risk stratification for ARDS after cardiac surgery. Using these predictors and ML models might provide a basis for early diagnostic and preventive strategies in the perioperative management of ARDS patients.

In vivo functional analysis of the cotton bollworm Helicoverpa armigera 24-dehydrocholesterol reductase (HaDHCR24) in phytosterol metabolism.

Insects have to obtain sterols from food due to the inability to synthesize this essential nutrient de novo. For lepidopteran insects, they can convert a variety of phytosterols into cholesterol to meet their growth needs. The final step of the cholesterol biosynthesis is the metabolism of desmosterol catalyzed by 24-dehydrocholesterol reductase (DHCR24). In this study, we identified a DHCR24 homolog in the cotton bollworm Helicoverpa armigera, designated as H. armigera 24-dehydrocholesterol reductase (HaDHCR24)-1. The quantitative expression analyses indicated that HaDHCR24-1 was highly enriched in the midgut where dietary sterol uptake occurs. Compared to the control, the DHCR24-1 mutant larvae generated by clustered regularly interspaced palindromic repeats (CRISPR) / CRISPR-associated nuclease 9 technology accumulated more desmosterol in the gut, while the content of cholesterol was significantly reduced. A similar phenomenon was observed when the DHCR24 inhibitor, amiodarone, was applied to the insects. Moreover, DHCR24-1 played an important role for the usage of ß-sitosterol, a major sterol in plants, in H. armigera, and loss of function of DHCR24-1 resulted in higher mortality on ß-sitosterol. However, the DHCR24 homolog does not necessarily exist in the genomes of all insects. The loss of this gene occurred more frequently in the insects feeding on animals, which further support the role of DHCR24-1 in using phytosterols. This gene may have important potential in developing new strategies to control herbivory pests in Lepidoptera and other insect orders.

Genetic Underpinnings of Cuticular Hydrocarbon Biosynthesis in the German Cockroach, Blattella germanica (L.): Progress and Perspectives.

Insect cuticular hydrocarbons (CHCs) serve as important waterproofing barriers and as signals and cues in chemical communication. Over the past 30 years, numerous studies on CHCs have been conducted in the German cockroach, Blattella germanica, leading to substantial progress in the field. However, there has not been a systematic review of CHC studies in this species in recent years. This review aims to provide a concise overview of the chemical composition, storage, transport, and physical properties of different CHCs in B. germanica. Additionally, we focus on the biosynthetic pathway and the genetic regulation of HC biosynthesis in this species. A considerable amount of biochemical evidence regarding the biosynthetic pathway of insect CHCs has been gathered from studies conducted in B. germanica. In recent years, there has also been an improved understanding of the molecular mechanisms that underlie CHC production in this insect. In this article, we summarize the biosynthesis of different classes of CHCs in B. germanica. Then, we review CHCs reaction to various environmental conditions and stressors and internal physiological states. Additionally, we review a body of work showing that in B. germanica, CHC profiles exhibit significant sexual dimorphism, specific CHCs act as essential precursors for female contact sex pheromone components, and we summarize the molecular regulatory mechanisms that underlie sexual dimorphism of CHC profiles. Finally, we highlight future directions and challenges in research on the biosynthesis and regulatory mechanisms of CHCs in B. germanica, and also identify potential applications of CHC studies in the pest control.

Two sex pheromone receptors for sexual communication in the American cockroach.

Volatile sex pheromones are vital for sexual communication between males and females. Females of the American cockroach, Periplaneta americana, produce and emit two sex pheromone components, periplanone-A (PA) and periplanone-B (PB). Although PB is the major sex attractant and can attract males, how it interacts with PA in regulating sexual behaviors is still unknown. In this study, we found that in male cockroaches, PA counteracted PB attraction. We identified two odorant receptors (ORs), OR53 and OR100, as PB/PA and PA receptors, respectively. OR53 and OR100 were predominantly expressed in the antennae of sexually mature males, and their expression levels were regulated by the sex differentiation pathway and nutrition-responsive signals. Cellular localization of OR53 and OR100 in male antennae further revealed that two types of sensilla coordinate a complex two-pheromone-two-receptor pathway in regulating cockroach sexual behaviors. These findings indicate distinct functions of the two sex pheromone components, identify their receptors and possible regulatory mechanisms underlying the male-specific and age-dependent sexual behaviors, and can guide novel strategies for pest management.

Haplotype-resolved 3D chromatin architecture of the hybrid pig.

In diploid mammals, allele-specific three-dimensional (3D) genome architecture may lead to imbalanced gene expression. Through ultradeep in situ Hi-C sequencing of three representative somatic tissues (liver, skeletal muscle, and brain) from hybrid pigs generated by reciprocal crosses of phenotypically and physiologically divergent Berkshire and Tibetan pigs, we uncover extensive chromatin reorganization between homologous chromosomes across multiple scales. Haplotype-based interrogation of multi-omic data revealed the tissue dependence of 3D chromatin conformation, suggesting that parent-of-origin-specific conformation may drive gene imprinting. We quantify the effects of genetic variations and histone modifications on allelic differences of long-range promoter-enhancer contacts, which likely contribute to the phenotypic differences between the parental pig breeds. We also observe the fine structure of somatically paired homologous chromosomes in the pig genome, which has a functional implication genome-wide. This work illustrates how allele-specific chromatin architecture facilitates concomitant shifts in allele-biased gene expression, as well as the possible consequential phenotypic changes in mammals.

Acetyl coenzyme A carboxylase modulates lipogenesis and sugar homeostasis in Blattella germanica.

Lipid and sugar homeostasis is critical for insect development and survival. In this study, we characterized an acetyl coenzyme A carboxylase gene in Blattella germanica (BgACC) that is involved in both lipogenesis and sugar homeostasis. We found that BgACC was dominantly expressed in the fat body and integument, and was significantly upregulated after molting. Knockdown of BgACC in 5th-instar nymphs did not affect their normal molting to the next nymphal stage, but it caused a lethal phenotype during adult emergence. BgACC-RNA interference (RNAi) significantly downregulated total free fatty acid (FFA) and triacylglycerol (TAG) levels, and also caused a significant decrease of cuticular hydrocarbons (CHCs). Repression of BgACC in adult females affected the development of oocytes and resulted in sterile females, but BgACC-RNAi did not affect the reproductive ability of males. Interestingly, knockdown of BgACC also changed the expression of insulin-like peptide genes (BgILPs), which mimicked a physiological state of high sugar uptake. In addition, BgACC was upregulated when B. germanica were fed on a high sucrose diet, and repression of BgACC upregulated the expression of the glycogen synthase gene (BgGlyS). Moreover, BgACC-RNAi increased the circulating sugar levels and glycogen storage, and a longevity assay suggested that BgACC was important for the survival of B. germanica under conditions of high sucrose uptake. Our results confirm that BgACC is involved in multiple lipid biogenesis and sugar homeostasis processes, which further modulates insect reproduction and sugar tolerance. This study benefits our understanding of the crosstalk between lipid and sugar metabolism.

Endovascular repair combined with in situ fenestration for new-onset or residual arch dissection in patients underwent aortic replacement.

INTRODUCTION: Ascending aorta or hemi-arch replacement is a frequently used treatment for patients with acute type A thoracic aortic dissection, particularly those who are elderly or have multiple comorbidities. However, in cases where there are secondary entry tears in the aortic arch or descending aorta, this procedure may not fully resolve the issue. The true lumen may remain compressed due to perfusion of the false lumen and usually require reoperation. METHODS: Between January 2019 and July 2022, 18 patients underwent endovascular total aortic arch repair and fenestration technique without requiring median re-sternotomy. Aortic stent grafts were implanted via the femoral approach, utilizing prosthetic vessels as an appropriate proximal landing zone for aortic stent graft deployment. Based on the debranching conditions of the arch in previous surgery, single, double or triple in situ fenestrations (ISFs) were performed, respectively. RESULTS: All 18 cases were technically successful, with a median follow-up period of 20 months (range: 18-31 months). All patients had a favourable postoperative course, with no deaths within 30 days or during their hospital stay. There were no instances of disabling stroke, paraplegia, endo-leak, stent graft migration or stent graft-induced new entry. In addition, all patients exhibited complete thrombosis of the false lumen at the level of the aortic arch. CONCLUSION: Our preliminary experience suggests that endovascular total arch repair combined with ISF technique is a viable, effective and safe option for treatment. Our mid-term results have been promising, but we acknowledge the need for further evaluation to assess long-term outcomes and durability.

A cell transcriptomic profile provides insights into adipocytes of porcine mammary gland across development.

BACKGROUND: Studying the composition and developmental mechanisms in mammary gland is crucial for healthy growth of newborns. The mammary gland is inherently heterogeneous, and its physiological function dependents on the gene expression of multiple cell types. Most studies focused on epithelial cells, disregarding the role of neighboring adipocytes. RESULTS: Here, we constructed the largest transcriptomic dataset of porcine mammary gland cells thus far. The dataset captured 126,829 high-quality nuclei from physiological mammary glands across five developmental stages (d 90 of gestation, G90; d 0 after lactation, L0; d 20 after lactation, L20; 2 d post natural involution, PI2; 7 d post natural involution, PI7). Seven cell types were identified, including epithelial cells, adipocytes, endothelial cells, fibroblasts cells, immune cells, myoepithelial cells and precursor cells. Our data indicate that mammary glands at different developmental stages have distinct phenotypic and transcriptional signatures. During late gestation (G90), the differentiation and proliferation of adipocytes were inhibited. Meanwhile, partly epithelial cells were completely differentiated. Pseudo-time analysis showed that epithelial cells undergo three stages to achieve lactation, including cellular differentiation, hormone sensing, and metabolic activation. During lactation (L0 and L20), adipocytes area accounts for less than 0.5% of mammary glands. To maintain their own survival, the adipocyte exhibited a poorly differentiated state and a proliferative capacity. Epithelial cells initiate lactation upon hormonal stimulation. After fulfilling lactation mission, their undergo physiological death under high intensity lactation. Interestingly, the physiological dead cells seem to be actively cleared by immune cells via CCL21-ACKR4 pathway. This biological process may be an important mechanism for maintaining homeostasis of the mammary gland. During natural involution (PI2 and PI7), epithelial cell populations dedifferentiate into mesenchymal stem cells to maintain the lactation potential of mammary glands for the next lactation cycle. CONCLUSION: The molecular mechanisms of dedifferentiation, proliferation and redifferentiation of adipocytes and epithelial cells were revealed from late pregnancy to natural involution. This cell transcriptomic profile constitutes an essential reference for future studies in the development and remodeling of the mammary gland at different stages.

Proteome Analysis of Male Accessory Gland Secretions in the Diamondback Moth, Plutella xylostella (Lepidoptera: Plutellidae).

In insects, male accessory gland proteins (ACPs) are important reproductive proteins secreted by male accessory glands (MAGs) of the internal male reproductive system. During mating, ACPs are transferred along with sperms inside female bodies and have a significant impact on the post-mating physiology changes of the females. Under sexual selection pressures, the ACPs exhibit remarkably rapid and divergent evolution and vary from species to species. The diamondback moth, Plutella xylostella (L.) (Lepidoptera: Plutellidae), is a major insect pest of cruciferous vegetables worldwide. Mating has a profound impact on the females' behavior and physiology in this species. It is still unclear what the ACPs are in this species. In this study, two different proteomic methods were used to identify ACPs in P. xylostella. The proteins of MAGs were compared immediately before and after mating by using a tandem mass tags (TMT) quantitative proteomic analysis. The proteomes of copulatory bursas (CB) in mated females shortly after mating were also analyzed by the shotgun LC-MS/MS technique. In total, we identified 123 putative secreted ACPs. Comparing P. xylostella with other four insect ACPs, trypsins were the only ACPs detected in all insect species. We also identified some new insect ACPs, including proteins with chitin binding Peritrophin-A domain, PMP-22/ EMP/ MP20/ Claudin tight junction domain-containing protein, netrin-1, type II inositol 1,4,5-trisphosphate 5-phosphatase, two spaetzles, allatostatin-CC, and cuticular protein. This is the first time that ACPs have been identified and analyzed in P. xylostella. Our results have provided an important list of putative secreted ACPs, and have set the stage for further exploration of the functions of these putative proteins in P. xylostella reproduction.

Construction of ultrasound-responsive urokinase precise controlled-release nanoliposome applied for thrombolysis.

Urokinase is widely used in the dissolution of an acute pulmonary embolism due to its high biocatalytic effect. However, how to precisely regulate its dose, avoid the side effects of hemolysis or ineffective thrombolysis caused by too high or too low a dose, and seize the golden time of acute pulmonary embolism are the key factors for its clinical promotion. Therefore, based on the precise design of a molecular structure, an ultrasonic-responsive nanoliposome capsule was prepared in this paper. Singlet oxygen is continuously generated under the interaction of the ultrasonic cavitation effect and the sonosensitizer protoporphyrin, and the generated singlet oxygen will break the thiol acetone bond between the hydrophilic head and the hydrophobic tail of the liposome, and the lipid The body structure disintegrates rapidly, and the urokinase encapsulated inside is rapidly released, down-regulating the expression of fibrinogen in the body, and exerting a thrombolytic function. The in vitro and in vivo results show that the smart urokinase nanoliposomes prepared by us have sensitive and responsive cytocompatibility to ultrasound and good in vivo thrombolytic properties for acute pulmonary embolism, which provides a new strategy for clinical acute pulmonary embolism thrombolysis.

A prodrug hydrogel with tumor microenvironment and near-infrared light dual-responsive action for synergistic cancer immunotherapy.

Immunotherapy has been used for cancer treatment, while it faces the common dilemmas of low therapeutic efficacy and serious immunotoxicity. In this study, we report the construction of a tumor microenvironment and near-infrared (NIR) light dual-responsive prodrug hydrogel for cancer synergistic immunotherapy in a more effective and safe manner. Such prodrug hydrogels were in-situ formed via calcium-induced gelation of alginate solution containing protoporphyrin IX (PpIX)-modified iron oxide (Fe3O4) nanoparticles and programmed death ligand 1 antibody (aPD-L1) prodrug nanoparticles crosslinked by reactive oxygen species (ROS)-responsive linkers. PpIX served as a photosensitizer to produce singlet oxygen (1O2) under NIR laser irradiation for photodynamic therapy (PDT), and Fe3O4 nanoparticles mediated chemodynamic therapy (CDT) to generate hydroxyl radical (·OH) via Fenton reaction in the tumor microenvironment. In view of the cumulative actions of PDT and CDT, amplified ROS was generated to not only induce immunogenic cell death (ICD), but also destroy ROS-responsive linkers to achieve on-demand release of aPD-L1 from prodrug nanoparticles. Boosted antitumor immunity was elicited in tumor-bearing mice due to the aPD-L1-mediated immune checkpoint blocking. As a result, the prodrug hydrogel-based synergistic immunotherapy could almost treat bilateral tumors and prevent lung and liver metastasis using 4T1 tumor mouse models. This study thus offers a dual-responsive prodrug hydrogel platform for precision cancer immunotherapy. STATEMENT OF SIGNIFICANCE: Via calcium-induced gelation of alginate, we constructed a prodrug hydrogel with tumor microenvironment and near-infrared light dual-responsive action for synergistic cancer immunotherapy. Such hydrogels can achieve on-demand release of aPD-L1 upon photoactivation in the tumor microenvironment. Through mediating photodynamic and chemodynamic therapy, the prodrug hydrogels can induce enhanced immunogenic cell death and synergistically improve the efficacy of aPD-L1-mediated immune checkpoint blocking. The prodrug hydrogel-based synergistic therapy almost deracinates the primary and distant tumors, and prevents lung and liver metastasis in tumor mouse models.

A single gene integrates sex and hormone regulators into sexual attractiveness.

Sex differentiation and hormones are essential for the development of sexual signals in animals, and the regulation of sexual signals involves complex gene networks. However, it is unknown whether a core gene is able to connect the upstream regulators for controlling sexual signal outputs and behavioural consequences. Here, we identify a single gene that integrates both sex differentiation and hormone signalling with sexual attractiveness in an insect model. CYP4PC1 in the German cockroach, Blattella germanica, controls the rate-limiting step in producing female-specific contact sex pheromone (CSP) that stimulates male courtship. As revealed by behavioural, biochemical, molecular, genetic and bioinformatic approaches, in sexually mature females, CYP4PC1 expression and CSP production are coordinately induced by sex differentiation genes and juvenile hormone (JH) signalling. In adult males, direct inhibition of CYP4PC1 expression by doublesexM binding in gene promoter and lack of the gonadotropic hormone JH prevent CSP production, thus avoiding male-male attraction. By manipulating the upstream regulators, we show that wild-type males prefer to court cockroaches with higher CYP4PC1 expression and CSP production in a dose-dependent manner, regardless of their sex. These findings shed light on how sex-specific and high sexual attractiveness is conferred in insects.

MiR-375-3p Promotes Cardiac Fibrosis by Regulating the Ferroptosis Mediated by GPX4.

Although coronary artery recanalization after myocardial infarction improves patient outcomes, inadequate ventricular remodeling following ischemia-reperfusion (IR) injury and secondary cardiac fibrosis (CF) are common and can lead to heart failure. MicroRNAs (miRNAs) play an important role in cardiovascular disorders. However, the underlying molecular mechanism of miRNAs in the occurrence and progression of CF has not been fully elucidated. Herein, through the construction of an I/R rat model and an angiotensin II-induced CF cell model, we evaluated the role of miR-375-3p in the progression of CF. In the I/R rat model and CF cell model, miR-375-3p promoted fibrosis by accelerating the ferroptosis of cardiomyocytes through mediating glutathione peroxidase 4 (GPX4). Furthermore, we treated the rats or cell model with miR-375-3p antagomir (or inhibitor) and ferroptosis inhibitor Ferrostatin-1 (Fer-1). The results showed that miR-375-3p antagomir (or inhibitor) and Fer-1 promoted the antioxidant capacity of cardiac fibroblasts, reduced GPX4-mediated ferroptosis process and alleviated I/R-induced CF. In conclusion, this study revealed that miR-375-3p directly targeted GPX4-an inhibitor of the ferroptosis pathway. Meanwhile, miR-375-3p can be a new potential biomarker for the prevention and treatment of CF.

Melanin synthesis genes BgTH and BgDdc affect body color and cuticle permeability in Blattella germanica.

Melanin is involved in cuticle pigmentation and sclerotization of insects, which is critical for maintaining structural integrity and functional completeness of insect cuticle. The 2 key enzymes of tyrosine hydroxylase (TH) and dopa decarboxylase (DDC) predicted in melanin biosynthesis are usually conserved in insects. However, it is unclear whether their function is related to epidermal permeability. In this study, we identified and cloned the gene sequences of BgTH and BgDdc from Blattella germanica, and revealed that they both showed a high expression at the molting, and BgTH was abundant in the head and integument while BgDdc was expressed highest in the fat body. Using RNA interference (RNAi), we found that knockdown of BgTH caused molting obstacles in some cockroaches, with the survivors showing pale color and softer integuments, while knockdown of BgDdc was viable and generated an abnormal light brown body color. Desiccation assay showed that the dsBgTH-injected adults died earlier than control groups under a dry atmosphere, but dsBgDdc-injected cockroaches did not. In contrast, when dsRNA-treated cockroaches were reared under a high humidity condition, almost no cockroaches died in all treatments. Furthermore, with eosin Y staining assay, we found that BgTH-RNAi resulted in a higher cuticular permeability, and BgDdc-RNAi also caused slight dye penetration. These results demonstrate that BgTH and BgDdc function in body pigmentation and affect the waterproofing ability of the cuticle, and the reduction of cuticular permeability may be achieved through cuticle melanization.

Two putative fatty acid synthetic genes of BgFas3 and BgElo1 are responsible for respiratory waterproofing in Blattella germanica.

Water retention is critical for physiological homeostasis and survival in terrestrial insects. While deposition of hydrocarbons on insect cuticles as a key measure for water conservation has been extensively investigated, we know little about other mechanisms for preventing water loss in insects. Here, we report two fatty acid synthetic genes that are independent of hydrocarbon production but crucial for water retention in the German cockroach Blattella germanica (L.). First, an integument enriched fatty acid elongase gene (BgElo1) was identified as a critical gene for desiccation resistance in B. germanica; however, knockdown of BgElo1 surprisingly failed to cause a decline in cuticular lipids. In addition, RNA interference (RNAi)-knockdown of an upstream fatty acid synthase gene (BgFas3) showed a similar phenotype, and transmission electron microscopy analysis revealed that BgFas3- or BgElo1-RNAi did not affect cuticle architecture. Bodyweight loss test showed that repression of BgFas3 and BgElo1 significantly increased the weight loss rate, but the difference disappeared when the respiration was closed by freeze killing the cockroaches. A water immersion test was performed, and we found that BgFas3- and BgElo1-RNAi made it difficult for cockroaches to recover from drowning, which was supported by the upregulation of hypoxia-related genes after a 10-h recovery from drowning. Moreover, a dyeing assay with water-soluble Eosin Y showed that this was caused by the entry of water into the respiratory system. Our research suggests that BgFas3 and BgElo1 are required for both inward and outward waterproofing of the respiratory system. This study benefits the understanding of water retention mechanisms in insects.

Downregulation of NADPH-cytochrome P450 reductase via RNA interference increases the susceptibility of Acyrthosiphon pisum to desiccation and insecticides.

Nicotinamide adenine dinucleotide phosphate (NADPH)-cytochrome P450 reductase (CPR) is involved in the metabolism of endogenous and exogenous substances, and detoxification of insecticides. RNA interference (RNAi) of CPR in certain insects causes developmental defects and enhanced susceptibility to insecticides. However, the CPR of Acyrthosiphon pisum has not been characterized, and its function is still not understood. In this study, we investigated the biochemical functions of A. pisum CPR (ApCPR). ApCPR was found to be transcribed in all developmental stages and was abundant in the embryo stage, and in the gut, head, and abdominal cuticle. After optimizing the dose and silencing duration of RNAi for downregulating ApCPR, we found that ApCPR suppression resulted in a significant decrease in the production of cuticular and internal hydrocarbon contents, and of cuticular waxy coatings. Deficiency in cuticular hydrocarbons (CHCs) decreased the survival rate of A. pisum under desiccation stress and increased its susceptibility to contact insecticides. Moreover, desiccation stress induced a significant increase in ApCPR mRNA levels. We further confirmed that ApCPR participates in CHC production. These results indicate that ApCPR modulates CHC production, desiccation tolerance, and insecticide susceptibility in A. pisum, and presents a novel target for pest control.

Genome-Wide Association Study Candidate Genes on Mammary System-Related Teat-Shape Conformation Traits in Chinese Holstein Cattle.

In the dairy industry, mammary system traits are economically important for dairy animals, and it is important to explain their fundamental genetic architecture in Holstein cattle. Good and stable mammary system-related teat traits are essential for producer profitability in animal fitness and in the safety of dairy production. In this study, we conducted a genome-wide association study on three traits-anterior teat position (ATP), posterior teat position (PTP), and front teat length (FTL)-in which the FarmCPU method was used for association analyses. Phenotypic data were collected from 1000 Chinese Holstein cattle, and the GeneSeek Genomic Profiler Bovine 100K single-nucleotide polymorphisms (SNP) chip was used for cattle genotyping data. After the quality control process, 984 individual cattle and 84,406 SNPs remained for GWAS work analysis. Nine SNPs were detected significantly associated with mammary-system-related teat traits after a Bonferroni correction (p < 5.92 × 10-7), and genes within a region of 200 kb upstream or downstream of these SNPs were performed bioinformatics analysis. A total of 36 gene ontology (GO) terms and 3 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were significantly enriched (p < 0.05), and these terms and pathways are mainly related to metabolic processes, immune response, and cellular and amino acid catabolic processes. Eleven genes including MMS22L, E2F8, CSRP3, CDH11, PEX26, HAL, TAMM41, HIVEP3, SBF2, MYO16 and STXBP6 were selected as candidate genes that might play roles in the teat traits of cows. These results identify SNPs and candidate genes that give helpful biological information for the genetic architecture of these teat traits, thus contributing to the dairy production, health, and genetic selection of Chinese Holstein cattle.

Impacts of Constitutive and Induced Benzoxazinoids Levels on Wheat Resistance to the Grain Aphid (Sitobion avenae).

Benzoxazinoids are important secondary metabolites in gramineae plants and have inhibitory and toxic effects against a wide range of herbivore pests. However, the relationship between benzoxazinoid level and plant resistance to aphids remains controversial. In this study, we investigated the relationship between benzoxazinoids composition and concentration in wheat leaves and the resistance to the grain aphid Sitobion avenae. Overall, six benzoxazinoids were detected and identified by mass spectrometry based metabolites profiling, including three lactams, two hydroxamic acids, and one methyl derivative. The constitutive levels of these benzoxazinoids were significantly different among the wheat varieties/lines. However, none of these benzoxazinoids exhibited considerable correlation with aphid resistance. S. avenae feeding elevated the level of 2-O-ß-D-glucopyranosyloxy-4,7-dimethoxy-(2H)-1,4-benzoxazin-3(4H)-one (HDMBOA-Glc) and reduced the level of 2-O-ß-D-glucopyranosyloxy-4-hydroxy-7-(2H)-methoxy-1,4-benzoxazin-3(4H)-one (DIMBOA-Glc) in some of the wheat varieties/lines. Moreover, aphid-induced level of DIMBOA-Glc was positively related with callose deposition, which was closely associated with aphid resistance. Wheat leaves infiltrated with DIMBOA-Glc caused a noticeable increase of callose deposition and the effect was in a dose dependent manner. This study suggests that the constitutive level of benzoxazinoids has limited impact on S. avenae. Aphid feeding can affect the balance of benzoxazinoids metabolism and the dynamic level of benzoxazinoids can act as a signal of callose deposition for S. avenae resistance. This study will extend our understanding of aphid-wheat interaction and provides new insights in aphid-resistance wheat breeding.